Nutrient modification of proliferation and radiation response in EMT6/Ro spheroids

The effects of external nutrients on the growth and radiation response of EMT6/Ro spheroids were studied by maintaining spheroids in media with different concentrations of glucose, amino acids, and vitamins. Compared to spheroids grown in normal glucose concentration (5.5 mM), spheroids grown in higher glucose media (24.8 mM), demonstrated no difference in initial volume doubling time, clonogenicity, number of proliferating cells, or cell cycle distributions. However, histology sections revealed that, spheroids grown in higher glucose concentration had a thicker viable rim than spheroids grown in normal glucose media. Two-step acridine orange staining and dual parameter flow cytometric analysis, in addition to continuous [3H]-thymidine labeling techniques, showed that spheroids grown in higher glucose had 2 to 3 times the fraction of quiescent cells, when compared to normal glucose spheroids. When irradiated in ice to reoxygenate, the Do's were similar in the normal and the higher glucose spheroids, but the Dq's were reduced in the higher glucose spheroids in the presence of increased amino acids and vitamins. When irradiated in air at 37 degrees C, spheroids grown in the higher glucose media were more sensitive (decreased Do), and had a smaller hypoxic fraction than when grown in normal glucose media. For spheroids grown in the same glucose concentrations but increased concentrations of amino acids and vitamins, there was generally an increased Do under all irradiation conditions. Some of these differences in radiation sensitivity could be correlated to differences in cellular glutathione levels of these spheroid cells.